Journal: Angiogenesis

Article Title: A human endothelial and adipose stem cell-based co-culture model for venous malformations

doi: 10.1007/s10456-026-10045-9

Figure Lengend Snippet: Transcriptomic analysis of hASC/HUVEC co-cultures. HUVECs were retrovirally transduced with bicistronic TIE2-IRES-GFP or GFP-expressing retroviruses and co-cultured with hASCs for 7 days. HUVECs (GFP + ) and hASCs (GFP - ) were separated by FACS and transcriptomic analysis was performed as indicated in Material and Methods. A - C Principal component analysis. Note the distinct clustering of genotypically different HUVECs and co-cultured hASCs. D and E Venn diagrams of the top 1000 up- and downregulated genes. Genes with adj. P < 0.05; Log2FC < –1 were classified as downregulated, while those with adj. P < 0.05; Log2FC > 1 were classified as upregulated. F Selected dot plots of the top 20 enrich pathways in hASC and in HUVECs ( TIE2 L914F vs. TIE2 WT ) potentially relevant to vascular anomalies. G Selected bar plots of the top 20 GO enrichment analyses for biological processes, cellular components and molecular functions in hASCs and HUVECs. All P-values < 1×10 -9 . The full GO and KEGG tables are shown in the supplementary material. H Heatmap of individual DEGs in HUVECs. The DEGs were selected based on the GO and KEGG analyses, potential involvement in VMs and biological function in ECs based on published data

Article Snippet: HUVECs were then cultured in ECGM2 (PromoCell, C-22011) supplemented with 10% fetal bovine serum (FBS, ScienCell, 0010), with the manufacturer’s supplement Mix solution (PromoCell, C-39215) and 1% penicillin/streptomycin (P/S; Sigma-Aldrich, P4333).

Techniques: Transduction, Expressing, Cell Culture

Journal: Angiogenesis

Article Title: A human endothelial and adipose stem cell-based co-culture model for venous malformations

doi: 10.1007/s10456-026-10045-9

Figure Lengend Snippet: vSMC tdT /HUVEC co-cultures. A Experimental scheme. Three genotypes of HUVECs, (HUVEC GFP , HUVEC GFP TIE2 WT and HUVEC GFP TIE2 L914F ), were cultured for 2 days after seeding vSMC tdTomato (vSMC tdT ) on top of the HUVEC layer, time-lapse imaged, or FACS sorted followed by RNA extraction and sequencing. B Migration velocity as quantified from time-lapse movies using a cell tracker. Means ± SD. * P < 0.05; ns, statistically non-significant in one way ANOVA followed by Tukey's post hoc test. n, three independent experiments. C and D Cellular morphology of vSMCs tdT in co-cultures of HUVEC and vSMC imaged after 24 h and 48 h. Genotypes of HUVECs are indicated on the left. The vSMCs tdT have a more elongated and narrower phenotype when cultured on top of HUVEC GFP TIE2 L914F . Scale bar, 50 µm

Article Snippet: HUVECs were then cultured in ECGM2 (PromoCell, C-22011) supplemented with 10% fetal bovine serum (FBS, ScienCell, 0010), with the manufacturer’s supplement Mix solution (PromoCell, C-39215) and 1% penicillin/streptomycin (P/S; Sigma-Aldrich, P4333).

Techniques: Cell Culture, RNA Extraction, Sequencing, Migration

Journal: Angiogenesis

Article Title: A human endothelial and adipose stem cell-based co-culture model for venous malformations

doi: 10.1007/s10456-026-10045-9

Figure Lengend Snippet: Transcriptomic analysis of vSMC/HUVEC co-cultures. A The PCA plot illustrates the distinct clustering of vSMCs co-cultured with the TIE2 genotypes indicated and harvested on days 2 and 4. B The volcano plots highlight the differential expression of vSMCs genes on days 2 and 4. The DEGs in blue/red are down/upregulated and the number of DEGs in each case is in dashed boxes. C Dot plot of potentially VM-related pathways among the top 20 KEGG-enriched pathways (vSMCs co-cultured with TIE2 L914F vs. TIE2 WT , at day 2). D Bar plot of GO enrichment analyses highlighting potentially VM-related pathways among the top 20 GOs (vSMCs co-cultured with TIE2 L914F vs. TIE2 WT at day 2). All P-values <1×10 -6 . The full GO and KEGG tables are shown in the supplementary data. E Heatmap of DEGs involved in vSMC contraction, ECM, cytoskeleton and F Notch signaling (at day 4)

Article Snippet: HUVECs were then cultured in ECGM2 (PromoCell, C-22011) supplemented with 10% fetal bovine serum (FBS, ScienCell, 0010), with the manufacturer’s supplement Mix solution (PromoCell, C-39215) and 1% penicillin/streptomycin (P/S; Sigma-Aldrich, P4333).

Techniques: Cell Culture, Quantitative Proteomics

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Setting. Dynamic in-vitro culture system. Original photos of the equipment and colorimetric map of the shear stress distribution: (A) - the peristaltic pump (Live Flow) able to perform an adjustable flow rate (100–450 μl/min) is connected both to bioreactor and reservoir mimicking blood flow circulation (left). Details of the LB1 (right, up) and magnification of the HUVEC monolayer inside the LB1 (right, bottom) are shown. (B) - colorimetric map of the shear stress distribution with wall shear stress magnitude diagrams. Up: the map illustrates the distribution of shear stress across the surface of the slide inside the LB1. The velocity is also indicated along the streamlines. Bottom: Shear stress distribution. The graph illustrates the shear stress distribution across the diameter of the LB1 slide, measured in the direction perpendicular to the inlet-outlet axis. The x-axis represents the slide’s length in tenths of a millimetre, and the y-axis shows the shear stress measured in Pascals (1 Pa = 10 dyne/cm 2 ). (C) - Dynamic device with Live Pa (left); the LB1connected to Live-Pa with the black piston inducing pressure increase (center); and the dynamic device equipped with two LB1 and Live-Pa inside the incubator (right).

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: In Vitro, Shear

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Experimental Design. Human vein umbilical endothelial cell (HUVEC) cultures in static (step 1) and dynamic conditions (step 2). Confluent cells were treated with ANG II for 24 h in both conditions. In step 2, 50% pressure increase was applied by Live-Pa to the circuit for 2 h, alone or in combination with ANG II.

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques:

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Intra-cellular signaling pathways in unstimulated or LPS-stimulated HUVEC seeded in LB1, cultured in static or dynamic conditions. NFkB (A) and p38MAPK (B) phosphorylation after stimulation with LPS (1 μg/mL). Endothelial cells seeded in LB1 for 24 h were incubated with LPS or medium alone both in static and under flow for further 24 h, as experimental control. Results are expressed as the ratio of phosphorylated to non-phosphorylated form, normalized to the respective control and evaluated using Image J software. Western Blotting images are representative of a single experiment. Histograms represent mean of three independent experiments. pNFkB: phosphorylated NFkB; p38MAPK: phosphorylated p38MAPK.

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: Protein-Protein interactions, Cell Culture, Phospho-proteomics, Incubation, Control, Software, Western Blot

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Intra-cellular signaling pathways in unstimulated or Angiotensin II-stimulated HUVEC seeded in LB1, cultured in static or dynamic conditions. NFkB (A) and p38MAPK (B) phosphorylation after stimulation with ANG II (1,000 nM). Endothelial cells seeded in LB1 for 24 h were incubated with Angiotensin II or medium alone both in static and under flow for further 24 h. Results are expressed as the ratio of phosphorylated to non-phosphorylated form, normalized to the respective control and evaluated using Image J software. Western Blotting images are representative of a single experiment. Histograms represent mean of three independent experiments. pNFkB: phosphorylated NFkB; p38MAPK: phosphorylated p38MAPK.

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: Protein-Protein interactions, Cell Culture, Phospho-proteomics, Incubation, Control, Software, Western Blot

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Intra-cellular signaling pathways in HUVEC seeded in LB1, cultured in static or dynamic conditions, unstimulated or stimulated with Angiotensin II and Live-PA, individually or in combination. NFkB (A) and p38MAPK (B) phosphorylation after stimulation with ANG II (1,000 nM) alone or in combination with Live-Pa. Endothelial cells seeded in LB1 for 24 h were stimulated with ANG II under flow for 24 h. Live-Pa was applied for further 2 h in the presence of medium or ANG II. Results are expressed as the ratio of phosphorylated to non-phosphorylated form, normalized to the dynamic control and evaluated using Image J software. Western Blotting images are representative of a single experiment. Histograms represent mean of three independent experiments. pNFkB: phosphorylated NFkB; p38MAPK: phosphorylated p38MAPK.

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: Protein-Protein interactions, Cell Culture, Phospho-proteomics, Control, Software, Western Blot

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Cytokine secretion. IL-6 (A) and IL-8 (B) protein levels in supernatants from HUVEC resting or stimulated with LPS (1 μg/mL) both in static and under flow for 24 h, as positive experimental control. Histograms represent mean +standard error of the mean (SEM). *p < 0.05 **p < 0.01 versus respective controls (static or dynamic medium). IL-6 (C) and IL-8 (D) protein levels in supernatants from HUVEC resting or stimulated with ANG II (1,000 nM) and Live-Pa, individually or in combination. Endothelial cells seeded in LB1 for 24 h were stimulated with ANG II (1,000 nM) under flow for 24 h. Live-Pa was applied for further 2 h in the presence of medium or ANG II. (Histograms represent mean +standard error of the mean (SEM) *p < 0.05 vs. medium + Live-Pa. **p < 0.01 versus medium (dynamic).

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: Control

Journal: Frontiers in Physiology

Article Title: Direct simulation of hypertensive stress on endothelial cells: a streamlined model of in-vitro-hypertension

doi: 10.3389/fphys.2025.1724932

Figure Lengend Snippet: Endothelin I secretion. (A) Endothelin I secretion in HUVEC seeded in LB1, cultured in static or dynamic conditions, and unstimulated or stimulated with LPS (1 μg/mL), as positive experimental control. Histograms represent mean +standard error of the mean (SEM). (B) Endothelin I secretion in HUVEC seeded in LB1, cultured in static or dynamic conditions, resting or in the presence of ANG II (1,000 nM) and Live-Pa, individually or in combination. Endothelial cells seeded in LB1 for 24 h were stimulated with ANG II (1,000 nM) under flow for 24 h. Live-Pa was applied for further 2 h in the presence of medium or ANG II. Histograms represent mean +standard error of the mean (SEM).

Article Snippet: Human umbilical vein endothelial cells (HUVEC) were purchased from Promocell (Heidelberg, Germany) and grown in the provided Endothelial Cell Growth Medium Ready-to-use (Promocell) in humidified incubator at 37 °C and 5% CO 2 .

Techniques: Cell Culture, Control